POS0348 ACTIVATING STAT3 MUTATIONS IN CD8+ T CELLS CORRELATE TO SEROLOGICAL POSITIVITY IN RHEUMATOID ARTHRITIS PATIENTS
Authors: K. Moosic, T. Olson, M. Freijat, S. Khalique, C. Hamele, B. Shemo, J. Boodoo, W. Baker, M. Schmachtenberg, A. Ratan, E. Darrah, F. Andrade, K. Olson, D. Feith, T. Loughran, D. Kimpel
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease that leads to inflammation and bone erosion of synovial joints. Serological markers such as rheumatoid factor (RF) and anti-CCP antibodies are commonly elevated, and cytotoxic CD8+ T-cell expansions have been implicated in disease pathogenesis. Large granular lymphocyte (LGL) leukemia is a rare hematologic malignancy characterized by clonal expansion of CD8+ cytotoxic T-cells. A major hallmark of all LGL leukemia is increased <i>STAT3</i> activation. About 50% of patients have somatic activating mutations in <i>STAT3</i>, the most common being Y640F and D661Y. Additionally, <i>STAT3</i> mutations correlate with numerous clinical features in LGL leukemia. LGL leukemia patients with <i>STAT3</i> mutations are more likely to respond to methotrexate treatment, tend to have low neutrophil counts, and clinical features can differ between mutation types (e.g. Y640F vs D661Y). Approximately one third of LGL leukemia patients also suffer from RA, and those with <i>STAT3</i> mutations are more likely to exhibit concomitant RA. Those patients with both diseases also tend to have increased positivity for serological markers like RF and anti-CCP antibodies than patients with either disease alone[1]. <h3>Objectives</h3> Based on disease overlap between LGL leukemia and RA, and a putative role for CD8+ T-cells in RA, we hypothesized that <i>STAT3</i> mutations may be detected in the CD8+ T cells of RA patients without LGL leukemia and correlate with RA clinical characteristics. <h3>Methods</h3> Blood samples, clinical parameters, and demographic data were collected from RA patients (n=96), many of whom were on immunosuppressives such as methotrexate, and healthy controls (n=9). CD8+ cell DNA was isolated and analyzed via droplet digital (dd)PCR to detect common <i>STAT3</i> mutations in LGL leukemia: Y640F, D661Y, and mutations in the S614 to G618 region. RA samples were divided into wild-type (WT) vs. <i>STAT3</i>-mutant groups using a variant allele frequency (VAF) cutoff of 0.05%, and clinical parameters were correlated to mutation status. <h3>Results</h3> Overall, RA patients had statistically different levels of STAT3 mutations compared to controls (D661Y p=0.0005, Y640F p=0.0185). In particular, many more low level (0.005-0.05% VAF) mutations were detected in the RA population (43/96 Y640F, 59/96 D661Y) vs. healthy controls (0/9 Y640F, 0/9 D661Y). Additionally, 3/96 RA samples had a <i>STAT3</i> mutation at a VAF (&gt;5%) large enough to be indicative of CD8+ T cell clonality compared to 0/96 controls (Table 1). Multiple serological RA markers were more frequently positive in <i>STAT3</i> mutant (80%-91%) relative to WT RA samples (44%-55%) (RF p=0.0005, anti-CCP p=0.018, double positivity p=0.006), with overall RF levels also significantly elevated (p=0.008) in <i>STAT3</i> mutant RA samples (Figure 1). Anti-CCP positivity was more common in Y640F patients (9/9) than D661Y (7/12). <h3>Conclusion</h3> Taken together, the presence of <i>STAT3</i> activating mutations in CD8+ T-cells and the association of mutations with sero-positivity suggest that <i>STAT3</i> activating mutations may play a role in the pathogenesis and clinical features in a subset of patients with RA. Further study will determine the impacts of <i>STAT3</i> mutation presence on disease severity, prognosis, and selection of treatment options/potential treatment responses. <h3>Reference</h3> [1]Moosic KB, Ananth K, Andrade F, Feith DJ, Darrah E, Loughran TPJ. Intersection Between Large Granular Lymphocyte Leukemia and Rheumatoid Arthritis. <i>Front Oncol</i> (2022) <b>0</b>:2083. doi: 10.3389/FONC.2022.869205 <h3>Acknowledgements</h3> The authors thank University of Virginia LGL Leukemia Registry personnel, especially Bryna Shemo and Andrea Hines, for their support of this study. We also thank Marieke Jones for her advice and time which were crucial to the statistical analysis of this study. <h3>Disclosure of Interests</h3> Katharine Moosic: None declared, Thomas Olson: None declared, Mark Freijat: None declared, Samara Khalique: None declared, Cait Hamele: None declared, Bryna Shemo: None declared, Jesse Boodoo: None declared, William Baker: None declared, Matthew Schmachtenberg: None declared, Aakrosh Ratan: None declared, Erika Darrah: None declared, Felipe Andrade: None declared, Kristine Olson: None declared, David Feith Grant/research support from: DJF has received research funding, honoraria and/or stock options from AstraZeneca, Dren Bio, Recludix Pharma and Kymera Therapeutics., Thomas Loughran Consultant of: TPL has received Scientific Advisory Board membership, consultancy fees, honoria and/or stock options from Keystone Nano, Bioniz Therapeutics, Dren Bio, Recludix Pharma, Kymera Therapeutics and Prime Genomics., Donald Kimpel Consultant of: DJK is on the Medical Advisory Board for Aurinia Pharmaceuticals and the Clinical Advisory Board for Landos Biopharma.